Because of the increasing use of nucleic acids in animal and human medicine, such as in gene therapy, there has been a growing need for making these materials available in greater purity. The term "nucleic acids" is to be understood as a collective term, encompassing deoxyribonucleic acids (DNA), ribonucleic acids (RNA), antisense RNA and nucleic acids with modified bases or structure. Endotoxins from gram-negative bacteria such as E. coli are often present in nucleic acids and when, during the course of recovery and isolation of such acids, particles of bacterial cell walls are not completely eliminated, there exists the possibility of the endotoxins finding their way to the final nucleic acid solutions being used in, say, a medical study. The introduction of endotoxins into patients must, of course, be avoided at all costs, since, where humans and experimental animals are concerned, endotoxins cause serious symptoms of illness, including fever and inflammation of blood vessels, activate blood-clotting and stimulate the production of antigens by the immune system.
EP 0 853 123 A1 discloses a process for the purification of nucleic acids in aqueous solutions, in which the solution containing nucleic acid and not cleared of endotoxins is flowed tangentially over ultrafiltration membranes with exclusion limits of 1 to 1000 kilodaltons. When this is carried out, because of its large size, a molecule of nucleic acid should not pass the membrane. At the same time, substances of lesser molecular weight, such as endotoxins, migrate through the membrane and/or are adsorbed on the membrane. The disadvantages of such an ultrafiltration technique are the considerable cost of the necessary apparatus, the large amount of time consumed (by virtue of the relatively low filtration speed common to the use of ultrafiltration membranes), the requirement for using large volumes of liquid feed solution (making the procedure unsuitable for laboratory scale work) and the tendency for dissolved proteins to prematurely blind the membranes.
These disadvantages are overcome by the present invention, which provides a process which can be carried out quickly and simply for the purification of aqueous solutions of nucleic acids, by which a lasting reduction of the endotoxin content is achieved, and which is suitable for both large- and small-scale treatments.